Aptamers are small, single stranded biomolecules, typically oligonucleotides (either DNA or RNA) or peptides, that bind to a specific target molecule (e.g. a protein or small molecule such as a steroid). They can be considered analogous to antibodies in their specificity but, unlike antibodies, aptamers are relatively small molecules. Peptide-based aptamers are generally less than thirty residues long while nucleotide-based aptamers are typically less than one hundred residues long.
Conventionally, aptamers are discovered using the SELEX process (Systematic Evoluatoin of Legands by Exponential Enrichment). See U.S. Pat. No. 5,637,459 (Burke). In this process, a large olignocleotide library (1014 to 1015 different sequences) of random sequences is exposed to the target molecule. Those sequences that do not bond to the target are removed. Those sequences that do bond are eluted and subjected to polymerase chain reaction (PRC). This process can be repeated multiple times until the tightest bonding sequence is isolated.
The SELEX process is somewhat limited. In certain occasions, a target is located that fails to tightly bond to any aptamer in the library of random sequences. The process is also time consuming. Depending on the particular application, this can require anywhere from a few days to a few months to complete. Additionally, isolated aptamers will often need to be re-engineered to reduce their sequence length and impart additional favorable biological properties. It would be desirable to find other methods for determining an aptamer sequence that do not suffer from at least some of this shortcomings.
Some virtual screening approaches to aptamer selection have been contemplated. See, for example: Chushak Y and Stone M O Nucl. Acids Res. 37 (2009) e87; Kim Bioinformatics 23: 2959-2960; Kim RNA 13: 478-492; Kevine Comput. Biol. and Chem. 31: 11-35. Over the past several years, various research groups have attempted to develop theoretical methods in order to speed up the SELEX process and design optimal aptamer sequences. Despite theoretical modeling of the working principle involved in SELEX process, various in silico approaches are favored by several groups. There are two general directions—a focus on developing a computational design of structured random pools such as RagPools and application of a virtual screening process.
Yet both directions possess some disadvantages. The first one is hindered by the requirement of a library of reasonably good starting sequences. It also requires prediction tools to arrive at candidate structures and the associated structural distributions. While this direction provides optimal choices for the design of a random pool for SELEX-prepared structures, in the final aptamer selection one still needs to go through the SELEX process in order to identify the “right” aptamer. The second direction requires information about the binding interfaces, “good” prediction of the tertiary structures of nucleotide sequences, and the availability of immense computational resources. One has to screen at most 4N N-mer RNA/DNA sequences to be able to analyze sequence diversity and structural complexity. The required computational power would be prohibitively large to complete the screening process. Another challenge involves the prediction of tertiary structures for arbitrary RNA/DNA sequences. Although the development of the RNA/DNA 2D and 3D structural prediction methods is promising, their validation is still questionable.